The present invention relates to enhancer peptide sequences originally derived from various retroviral envelope (gp4l) protein sequences that enhance the pharmacokinetic properties of any core polypeptide to which they are linked. The invention is based on the discovery that hybrid polypeptides comprising the enhancer peptide sequences linked to a core polypepide possess enhanced pharmacokinetic properties such as increased half life. The invention further relates to methods for enhancing the pharmacokinetic properties of any core polypeptide through linkage of the enhancer peptide sequences to the core polypeptide. The core polypeptides to be used in the practice of the invention can include any pharmacologically useful peptide that can be used, for example, as a therapeutic or prophylactic reagent. In a non-limiting embodiment, the invention is demonstrated by way of example wherein a hybrid polypeptide comprising, for example, an HIV core polypeptide linked to enhancer peptide sequences, is shown to be a potent, non-cytotoxic inhibitor of HIV-1, HIV-2 and SIV infection. Additionally, the enhancer peptide sequences of the invention have been linked to a respiratory syncytial virus (RSV) core polypeptide and a luteinizing hormone receptor (LH-RH) core polypeptide. In each instance, the hybrid polypeptide was found to possess enhanced pharmacokinetic properties, and the RSV hybrid polypeptide exhibited substantial anti-RSV activity.
The Sequence Listing for this application is on duplicate compact discs labeled xe2x80x9cCopy 1xe2x80x9d and xe2x80x9cCopy 2xe2x80x9d. Copy 1 and Copy 2 each contain only one file named xe2x80x9c7872081.APPxe2x80x9d which was created on Aug. 10, 2001 and is 645,776 bytes. The entire contents of each of the compact discs is incorporated herein by reference.
Polypeptide products have a wide range of uses as therapeutic and/or prophylactic reagents for prevention and treatment of disease. Many polypeptides are able to regulate biochemical or physiological processes to either prevent disease or provide relief from symptoms associated with disease. For example, polypeptides such as viral or bacterial polypeptides have been utilized successfully as vaccines for prevention of pathological diseases. Additionally, peptides have been successfully utilized as therapeutic agents for treatment of disease symptoms. Such peptides fall into diverse categories such, for example, as hormones, enzymes, immunomodulators, serum proteins and cytokines.
For polypeptides to manifest their proper biological and therapeutic effect on the target sites, the polypeptides must be present in appropriate concentrations at the sites of action. In addition, their structural integrity must generally be maintained. Therefore, the formulation of polypeptides as drugs for therapeutic use is directed by the chemical nature and the characteristics of the polypeptides, such as their size and complexity, their conformational requirements and their often complicated stability, and solubility profiles. The pharmacokinetics of any particular therapeutic peptide is dependent on the bioavailability, distribution and clearance of said peptide.
Since many bioactive substances, such as peptides and proteins, are rapidly destroyed by the body, it is critical to develop effective systems for maintaining a steady concentration of peptide in blood circulation, to increase the efficacy of such peptides, and to minimize the incidence and severity of adverse side effects.
The present invention relates, first, to enhancer peptide sequences derived from various retroviral envelope (gp41) protein sequences i.e., HIV-1, HIV-2 and SIV, that enhance the pharmacokinetic properties of any core. polypeptide to which they are linked. The invention is based on the surprising result that when the disclosed enhancer peptide sequences are linked to any core polypeptide, the resulting hybrid polypeptide possesses enhanced pharmacokinetic properties including, for example, increased half life and reduced clearance rate relative to the core polypeptide alone. The present invention further relates to such hybrid polypeptides and core polypeptides.
The core polypeptides to be used in the practice of the invention can comprise any peptides which may be introduced into a living system, for example, any peptides capable of functioning as therapeutic or prophylactic reagents useful for treatment or prevention of disease. Such peptides include, for example, growth factors, hormones, cytokines, angiogenic growth factors, extracellular matrix polypeptides or polypeptides that exhibit antifusogenic and/or antiviral activity, and peptides or polypeptides that function as immunogens including, for example, viral and bacterial polypeptides.
The invention further relates to methods for enhancing the pharmacokinetic properties of any core polypeptide through linkage of the core polypeptide to the enhancer peptide sequences to form hybrid polypeptides.
The invention is demonstrated by way of examples wherein hybrid polypeptides containing an HIV core polypeptide linked to enhancer peptide sequences are shown to exhibit greatly enhanced pharmacokinetic properties and act as a potent, non-cytotoxic inhibitors of HIV-1, HIV-2 and SIV infection. The invention is further demonstrated by examples wherein hybrid polypeptides; containing an RSV core polypeptide or a luteinizing hormone polypeptide are shown to exhibit greatly enhanced pharmacokinetic properties. In addition, the RSV hybrid polypeptide exhibited substantial anti-RSV activity.
Peptides are defined herein as organic compounds comprising two or more aminoacids covalently joined by peptide bonds. Peptides may be referred to with respect to the number of constituent natural L-amino acids, i.e., a dipeptide contains two amino acid residues, a tripeptide contains three, etc. Peptides containing ten or fewer amino acids may be referred to as oligopeptides, while those with more than ten amino acid residues are polypeptides. Such peptides may also include non-natural amino acids and any of the modifications and additional amino and carboxyl groups as are described herein.
Peptide sequences defined herein are represented by one-letter symbols for amino acid residues as follows:
A (alanine)
R (arginine),
N (asparagine)
D (aspartic acid)
C (cysteine)
Q (glutamine)
E (glutamic acid)
G (glycine)
H (histidine)
I (isoleucine)
L (leucine)
K (lysine)
M (methionine)
F (phenylalanine)
P (proline)
S (serine)
T (threonine)
W (tryptophan)
Y (tyrosine)
V (valine)
X (any amino acid)
xe2x80x9cEnhancer peptide sequencesxe2x80x9d are defined as peptides having the following consensus amino acid sequences: xe2x80x9cWXXWXXXIxe2x80x9d, xe2x80x9cWXXWXXXxe2x80x9d, xe2x80x9cWXXWXXxe2x80x9d, xe2x80x9cWXXWXxe2x80x9d, xe2x80x9cWXXWxe2x80x9d, xe2x80x9cWXXXWXWXxe2x80x9d, xe2x80x9cXXXWXWXxe2x80x9d, xe2x80x9cXXWXWXxe2x80x9d, xe2x80x9cXWXWXxe2x80x9d, xe2x80x9cWXWXxe2x80x9d, xe2x80x9cWXXXWXWxe2x80x9d, xe2x80x9cWXXXWXxe2x80x9d, xe2x80x9cWXXXWxe2x80x9d, xe2x80x9cIXXXWXXWxe2x80x9d, xe2x80x9cXXXWXXWxe2x80x9d, xe2x80x9cXXWXXWxe2x80x9d, xe2x80x9cXWXXW xe2x80x9d, xe2x80x9cXWXWXXXWxe2x80x9d, xe2x80x9cXWXWXXXxe2x80x9d, xe2x80x9cXWXWXXxe2x80x9d, xe2x80x9cXWXWXxe2x80x9d, xe2x80x9cXWXWxe2x80x9d, xe2x80x9cWXWXXXWxe2x80x9d, or xe2x80x9cXWXXXWxe2x80x9d, wherein X can be any amino acid, W represents tryptophan and I represents isoleucine. As discussed below, the enhancer peptide sequences of the invention also include peptide sequences that are otherwise the same as the consensus amino acid sequences but contain amino acid substitutions, insertions or deletions but which do not abolish the ability of the peptide to enhance the pharmacokinetic properties of a core peptide to which it is linked.
xe2x80x9cCore polypeptidexe2x80x9d as used herein, refers to any polypeptide which may be introduced into a living system and, thus, represents a bioactive molecule, for example any polypeptide that can function as a pharmacologically useful peptide for treatment or prevention of disease.
xe2x80x9cHybrid polypeptidexe2x80x9d as used herein, refers to any polypeptide comprising a terminal enhancer peptide sequence and a core polypeptide.